Covalent modifications of nucleic acids have been used to explore the diverse functions of nucleic acids and to characterize their intrinsic biochemical properties in biological systems. Continuous development of covalent conjugation methods for nucleic acids further facilitates applications of nucleic acids as research tools for chemical and biomedical studies, improves potency, and promotes specificity of nucleic acids as therapeutic reagents in medicine. Oligonucleotides conjugated with peptides to afford peptide-oligonucleotide conjugates (POCs) hold promise as an effective therapeutic reagent to treat viral infections and genetic diseases. Applications of POCs in basic and clinical research have produced nucleic acids with improved biological stability, cellular uptake efficiency, and in vivo cell-specific targeting.
Preparation of POCs is essential for therapeutic applications of nucleic acids such as oligonucleotides. The direct use of oligonucleotides in medicine generally falls short of disease treatment expectations due to poor cell specificity and uptake of nucleic acids, and inaccessibility of nucleic acids to cell nuclei. The conjugation of oligonucleotides with peptides is thus the most common approach to circumvent cell delivery and specificity problems of oligonucleotides. However, methods currently available to prepare POCs are inefficient and inconvenient for typical research laboratories. The difficulties experienced when preparing desired effective POCs restrict the broad applications of POCs in achieving their much-needed medical applications. POCs are generally prepared by post solid-phase synthesis to couple peptides with oligonucleotides (fragment coupling strategy) or stepwise solid-phase synthesis. The generic fragment coupling strategy separately synthesizes peptides and oligonucleotides by standard solid-phase synthesis protocols, and then both are covalently linked together by post solid-phase synthesis. There are limited reactions available for POC formation because peptides primarily use unstable phosphodiester linkages when conjugating to 5′- or 3′-termini or ester linkages when linking to 2′-positions of riboses in oligonucleotides. The development of a facile approach to use hydroxyl or phosphate groups in standard oligonucleotides to afford stable POCs with high purity and yields is highly desirable.
In order to overcome the drawbacks in the prior art, a method for conjugating nucleic acids with small molecules is provided. The particular design in the present invention not only solves the problems described above, but also is easy to be implemented. Thus, the present invention has potential use in the industries.